Adsorptive processes and devices are widely used in the analysis and purification of chemicals, including synthetic and naturally-derived pharmaceuticals, blood products and recombinant proteins. Chromatography is a general separation technique that relies on the relative affinity or distribution of the molecules of interest between a stationary phase and a mobile phase for molecular separation. The stationary phase typically comprises a porous media imbibed with solvent. The mobile phase comprises a solvent, which can be aqueous or organic, that flows through the interstitial space that exists between the spaces occupied by the stationary phase.
Columns with associated end caps, fittings and tubing are the most common configuration, with the media packed into the tube or column. The mobile phase is pumped through the column. The sample is introduced at one end of the column, the feed end, and the various components interact with the stationary phase by any one of a multitude of adsorptive phenomena. The differential adsorptive interaction between the components and media leads them to traverse the column at different velocities, which results in a physical separation of the components in the mobile phase. The separated components are collected or detected at the other end of the column, the eluent end, in the order in which they travel in the mobile phase. In one type of adsorptive process, referred to as capture and release process, the process involves multiple steps, first to load the media, then to wash it, and then to elute it.
Chromatographic methods include among other methods, gel chromatography, ion exchange chromatography, hydrophobic interaction chromatography, reverse phase chromatography, affinity chromatography, immuno-adsorption chromatography, lectin affinity chromatography, ion affinity chromatography and other such well-known chromatographic methods.
Adsorptive media comes in many forms, most typically in the form of chromatographic beads. The beads are conventionally packed into columns, with the column walls and ends immobilizing the beads into a fixed adsorptive bed, a bed being a porous three dimensional structure containing the stationary phase (in this case the beads) and the pore space through which the mobile phase flows/permeates (the space between the beads).
Conventional chromatographic devices require that beads must be packed into a column. The quality of this packing determines the performance of the adsorbing bed. This adds another source of variability to the chromatographic process and must be validated before use. Furthermore, beds packed with beads are prone to voiding, a phenomenon whereby the beads settle into a denser structure resulting in the creation of voids and in non-homogeneities in the packing density of the bed, all of which results in a deterioration of performance. This is especially true in columns packed with soft beads.
While it is theoretically possible to achieve high operational velocities exceeding 2000 cm/hr with rigid particles, beads such as AbSolute (Novasep Process Pompey, France) or silica beads packed in columns, it is generally not possible to use softer beads such as agarose and polymethylmethacrylate microspheres (PMMA) at high velocities exceeding 400 cm/hr.
Furthermore silica particles are not generally used to purify monoclonal antibodies. Velocities higher than 400 cm/hr are even less possible with production-scale columns having diameters of 20-200 cm (Sigma=0.02-0.2/cm). Where Sigma is a specific surface area of the adsorptive bed, defined as the surface area of solid support divided by the volume of the bed, a measure of the level of support imparted to the packed bed by the walls of the adsorptive device. In the case of a conventional column sigma equals four divided by d (where d equals the diameter of the column).
Some silica bead based processes can theoretically be run at a velocity of 2000 cm/hr, however this is not possible with agarose or PMMA beads. It is also difficult to scale up lab processes to manufacturing volumes (e.g. exceeding one liter). However, even beds packed with rigid beads may be compressible, leading to undesirable increases in pressure drop and possibly voiding if run at very high velocities.